Table of Contents 21-3

cAMP-Response Element Binding Protein (CREB) and Gene Activation

Cyclic adenosine monophosphate (cAMP) serves as a second messenger in multiple cell signaling pathways. As well, cAMP stimulates protein kinase A-mediated phosphorylation of the cAMP-response element binding protein (CREB) family at a conserved serine (Ser-133 in CREB1), which in turn leads to the recruitment of coactivators, including CREB-binding protein (CBP)/p300, TORC and TAF_II4. Phospho-CREB (P-CREB) binds cAMP-response elements (CREs) in gene promoters, and thereby coordinates with other transcription factors to recruit RNA polymerase II to the promoters and stimulate transcription of target genes.

Zhang and colleagues used genome-wide approaches, including microarray analyses and chromatin immunoprecipitation (ChIP) assays, to characterize the target genes and regulation of CREB activity in different human tissues. They started by searching for full CREs (TGACGTCA) and half-CREs (TGACG/CGTCA) that were conserved in human, mouse, and rat genome alignments. Promoters were defined as 3kb upstream to 300 bp downstream of annotated transcription start sites; downstream (within 300 bp) TATA boxes were sought for all CREs that were identified in promoter sequences. In the human genome 10,447 CREs and 740,390 half-CREs were found, with the majority of conserved CREs occurring within 200bp of the transcription start site. Combining 3 independent algorithms, the authors identified 4 084 putative CREB target genes, of which 1 518 also contained TATA boxes. CREB target genes were mostly transcription factors (38%), followed by genes involved in metabolic control, cell-cycle regulation, and regulated secretion.

The authors proceeded to investigate mechanisms of regulation of CREB binding that may explain the tissue specificity of CREB target gene profiles. About 3000 promoters (17% of protein-coding genes) were found to be CREB occupied in vivo. CpG methylation at the CRE inhibits CREB binding, and some CREs were found to be always unmethylated (ie, CREB-responsive), some universally methylated (ie, silenced genes), and others methylated only in certain cell types. Levels of P-CREB were low in resting conditions, but increased markedly within one hour of exposure to forskolin, a cAMP agonist; changes in P-CREB levels did not differ between cAMP inducible versus noninducible genes, so differential CREB phosphorylation does not construe a mechanism for tissue specificity of target gene profiles. Forskolin induced appoximately 100 genes in 3 different cell types tested, though the sets were mostly distinct. Likewise, cAMP stimulated growth factor genes and antiapoptotic genes in cultured human islets, in contrast to primary hepatocytes, where it stimulated genes involved in fasting glucose and lipid metabolism. Recruitment of CBP and other CREB regulatory partners to promoters by cAMP or forskolin was another mechanism identified for tissue specificity of the target gene profiles.

Zhang X, Odom DT, Koo S-H, et al. Genome-wide analysis of cAMP-response element binding protein occupancy, phosphorylation, and target gene activation in human tissues. Proc Natl Acad Sci. 2005;102:4459-4464.

Editor’s Comment: The power of genome-wide bioinformatics makes it now feasible to study the response patterns to such fundamental signaling mechanisms like the cAMP pathway. I refer the reader to the databases posted by the authors as excellent reference tools. For example, at http://natural.salk.edu/CREB, one can search human, mouse and rat genes of interest to see if/where they have CREB binding and inducibility by cAMP in various tissues. cAMP is an important second messenger for multiple water-soluble hormones that bind cell surface receptors. Such hormones include: somatostatin, chorionic gonadotropin (HCG), lipotropin (LPH), melanocyte stimulating hormone (MSH), adrenocorticotrophic hormone (ACTH), follicle-stimulating hormone (FSH), luteinizing hormone (LH), thyroid stimulating hormone (TSH), antidiuretic hormone (ADH), parathyroid hormone (PTH), calcitonin and glucagon. Various reviews of cAMP signaling in the endocrine system are listed below.^1-7

Adda Grimberg, MD

References - (linked to )

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